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In nature, biomolecules guide the formation of hierarchically-ordered, lightweight, inorganic-organic composites such as corals, shells, teeth and bones. M13 bacteriophage has been used to mimic bio-inspired material development due to its rigid, nanoscale rod-like morphology. Liquid-crystalline monolayers of genetically engineered phage have been used to template crystallization of thin layers of inorganic and metallic materials. We have created thin films composed of engineered M13 phage capable of binding inorganic components. We employed both a dip-cast and a drop-cast film fabrication method on both smooth and rough gold, silica and glass casting surfaces to create thin films and 3D structures of various degrees of hierarchical order. We have found the engineered M13 phage and the inorganic mineral significantly affected both film morphology and the mechanical properties of the film. Similarly, film fabrication parameters such as solution chemistry, temperature, and pulling speed affected film properties. Using a calcium phosphate biomineralized 4E phage, film thickness increased linearly with the number of layers/dips in the phage solution. The stiffness of these composites (Young's modulus) were >80 GPa for mineralized, multilayer films. These materials are an order of magnitude stiffer than the biological equivalent collagen. Stiffness, however, does not appear to increase in a multilayer film beyond a saturation point. Ultimately, we have developed a platform for phage-based bio-composites for developing high performance materials.

Magnesium (Mg) plays an important role in the body mediating cell-extracellular matrix (ECM) interactions, bone apatite structure and density, and nucleic acid chemistries. While Mg has been investigated as a biomaterial for bone applications, it has not been studied for applications within soft tissues. This study investigated, for the first time, the response of fibroblasts to magnesium oxide (MgO) nanoparticles for soft tissue engineering applications. Primary human dermal fibroblasts were cultured both on tissue culture polystyrene in media supplemented with MgO nanoparticles as well as on poly-L-lactic acid (PLLA)-MgO nanoparticle composites. As this study was conducted concurrently with a study aimed at bone tissue engineering, hydroxyapatite (HA) nanomaterials were used for comparison. Results showed for the first time that fibroblasts adhered onto MgO-containing composites roughly three times better than HA-PLLA samples and roughly 4.5 times better than plain PLLA samples. Fibroblasts also proliferated to statistically higher densities when cultured in medium supplemented with MgO nanoparticles compared to un-supplemented medium and medium supplemented with HA nanoparticles. These preliminary results together suggest that MgO nanoparticles should be further investigated as materials to improve the regeneration of soft tissues as well as bone.

MscL, a large-conductance mechanosensitive channel, is a ubiquitous osmolyte release valve that aids bacteria in surviving abrupt hypo-osmotic shocks. The large scale of its tension-driven opening transition makes it a strong candidate to serve as a transducer in novel stimuli-responsive biomolecular materials. In the previous work, a low-threshold gain-of-function V23T mutant of MscL produced a reliable activation behavior in a droplet interface bilayer (DIB) with applied axial droplet compression. Near the maximal compression, the aqueous droplets deform and the resulting increase in surface area leads to an increase in tension in the water-lipid-oil interface. This increase in tension is the product of the relative change in the droplet surface area and the elastic modulus of the DPhPC lipid monolayer (∼120 mN/m). This paper, presents a study of the physical processes that cause MscL gating in the DIB. Analysis of video during compression and relaxation of the droplets is utilized to estimate the change in the surface area of the droplet and the variation on monolayer surface tension. The monolayer surface tension is proportional to the area change of the droplet normalized to the original surface area. The results demonstrate that the area change in the droplet is negligible at frequencies above 1 Hz, but is approximately 2% at frequencies in the range of 100 mHz. In addition, at low frequencies (∼0.2 Hz) bilayer thinning occurs at maximum compression, proving an increase in bilayer tension. However, this study also shows that gating at frequencies higher than 0.2 Hz could be achieved through the application of high duty cycle oscillation (∼75%). The relative change in monolayer area increases significantly at higher duty cycle oscillations where the compression stroke is much faster than the relaxation stroke.

Droplet interface bilayers (DIBs) are formed using brain total lipid extract (BTLE) to create a synthetic bilayer whose lipid composition mimics that of neural cells. The electrical properties of BTLE DIBs, specifically membrane resistance, capacitance, and rupture potential, are determined and compared to the properties of bilayers formed using DPhPC, the most common lipid within the growing DIB field. There is no significant difference in the resistance or rupture potential of BTLE and DPhPC bilayers, for instance with average nominal resistance over 200 GΩ and rupture potential around 200 mV. In electrical measurements with either DPhPC or BTLE bilayers, applied voltages of up to ±150 mV yield low levels of leakage current. Upon interaction with the pore-forming amyloid-beta (Aβ) peptide, both bilayers display sudden significant voltage-dependent increases in conductance with characteristic threshold voltages well below 150 mV. Discrete single-channel type events are observed in the case of Aβ-BTLE whereas disordered fluctuating conductance is observed with Aβ-DPhPC. Circular dichroism is measured for Aβ incubated with BTLE and DPhPC liposomes, as well as pure Aβ, at a range of temperatures over a period of several weeks. Changes in secondary structure of liposome-bound and pure Aβ are significantly affected by both lipid type and temperature. A key finding includes the 100% conversion of Aβ to alpha-helical confirmation within 24 hours when incubated with liposomes (of either type) at physiologically relevant 37°C. The 100% alpha-helical Aβ is maintained for up to 2 weeks at 37°C when incubated with liposomes, although other structures begin to emerge after the 14 day mark. Between 14-31 days after reconstitution, Aβ incubated at 37C with BTLE bilayers displays longer lasting alpha-helical content than DPhPC. At the same temperature, pure Aβ is 100% alpha helical only at the 1 day mark with apparent restructuring from day 2 through day 31. Refrigerated Ab samples do not display 100% alpha-helical structure across the entire 31 day testing period. The differences observed between BTLE and DPhPC in both electrophysiological and spectroscopic experiments may be a result of phase separations or other variations in membrane fluidity that result from the use of a homogeneous total lipid extract. Time and temperature play essential roles in the aggregation and restructuring of potentially toxic Aβ oligomers, and there is motivation for further efforts to elicit the mechanistic differences in interactions of Ab with BTLE compared to DPhPC.

Designing new materials with well-defined structures and desired functions is a challenge in materials science, especially with nanomaterials. Nature, however, solves design of these materials through a self-assembling, hierarchically ordered process. We have investigated the mechanisms by which the high- aspect ratio and unique surface chemistry of M13 bacteriophage can give rise to increasingly complex, hierarchically ordered, bundled phage structures with a wide range of material applications. A molecular dynamic simulation of the 3-D structure of a 20-nm section of wild type (WT) and mutant phage types were developed based on WT phage crystal structure and ab initio calculations. Simulations of these phage were then used to examine repulsive and attractive forces of the particles in solution. Examination of contact interactions between two WT phage indicated the phage were maximally attracted to each other in a head to tail orientation. A mutant phage (4E) with a higher negative surface charge relative to WT phage also preferentially ordered head to tail in solution. In contrast, a mutant phage (CLP8) with a net positive surface charge had minimal repulsion in a 90° orientation. Understanding the self-assembly process through molecular dynamic simulations and decomposition of fundamental forces driving inter- and intra-strand interactions has provided a qualitative assessment of mechanisms that lead to hierarchical phage bundle structures. Results from simulation agree with experimentally observed patterns from self-assembly. We anticipate using this system to further investigate development of hierarchical structures not only from biological molecules but also from synthetic materials.

Organic biological hybrid systems, accessible by covalent functionalization of photosynthetic proteins with molecular antennas, represent a promising novelty to enhance natural photosynthesis. In this paper, we present the successful bioconjugation of a commercial fluorophore, fluorescein isothiocyanate (FITC), to the photosynthetic reaction center RC from the bacterium Rhodobacter sphaeroides strain R26. The resulting hybrid outperforms the pristine protein in hole-electron couple generation yield, exclusively at wavelengths where the fluorophore absorbs while the protein does not.